Despite a robust antibody response to HIV following infection, the neutralizing antibody response is poor and ineffective at controlling disease;however, specific antibodies have been shown effective at preventing infection in non-human primate models. Thus, novel immunotherapeutic approaches are necessary for antibodies to be useful in treating HIV infection. In addition to the direct neutralization of virus by antibodies, there is a broader potential for antibody mediated inhibition of HIV by targeting HIV to effector cells. We hypothesize that directing HIV, using bispecific antibodies incorporating select anti-HIV antibodies and anti-IgA receptor (CD89) antibody, to neutrophils will effectively destroy HIV and HIV infected cells. Neutrophils, which are the most predominant type of white blood cells, are very efficient at mediating cell cytotoxicity as well as relatively resistant to infection with HIV. Therefore, they represent a significant arsenal against infection if they can be directed and armed to destroy HIV and infected cells. Based on structure-function relationships, we propose to study bispecific antibodies generated using three different human anti-HIV monoclonal antibodies. IgG1b12 is a neutralizing antibody with a unique structure giving it improved access to the CD4 binding site of virus. F425B4e8 is also a potent neutralizing antibody, to the V3 loop, which is relatively accessible on virus and infected cells. Finally, given the exposure of the well-conserved cluster I epitope on gp41, recognized by the antibody F240, we also propose to determine the ability of a bispecific antibody, with broad anti-HIV reactivity, to destroy HIV. The use of F240 allows clear differentiation between inherent neutralization activity from neutrophil directed killing. Bispecific antibodies will be tested for the ability to stimulate neutrophil effector functions and the destruction of HIV. Bispecific antibody constructs will be prepared as single chain Fv (scFv) antibodies. However, since the short half-life of scFv antibodies in vivo may be limiting, we also propose to generate constructs with a modified Fc domain. It is expected that some constructs will be more effective than others based in part on epitope exposure. Constructs will also be tested for bystander killing of uninfected cells with bound gp120. Identification of constructs active in vitro at inhibiting viral production, with minimal bystander killing, will allow for further testing in non-human primate models of infection. These studies represent an important advance in treating chronic HIV infection. D PUBLIC HEALTH RELEVANCE: espite the success of combination therapy with anti-retroviral agents, additional means of therapy are required for the treatment of HIV infection. Since antibodies that directly inhibit virus are either ineffective or rare in vivo, we propose to design antibodies to target HIV to cells for destruction. These studies represent an important advance in treating chronic HIV infection.